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The effect of dietary starch and fat content on serum creatine kinase (CK) activity and substrate availability was evaluated in 4 mares of Quarter Horse-related breeds with polysaccharide storage myopathy (PSSM). Four isocaloric diets ranging in digestible energy (DE) from 21.2% (diet A), 14.8% (B), 8.4% (C), to 3.9% (D) for starch, and 7.2% DE (diet A), 9.9% (B), to 12.7% DE (diet C and D) for fat were fed for 6-week periods (4 weeks with exercise) using a 4 X 4 Latin square design. Postprandial glucose and insulin responses were measured, and 4 hours postexercise, serum CK activity, glucose, insulin, free fatty acids (FFA), and beta-hydroxybutyrate (beta-HBA) were analyzed. Glycogen, glucose-6-phosphate, citrate synthase, 3-hydroxy-acyl-CoA dehydrogenase, lactate dehydrogenase as well as abnormal polysaccharide and lipid content were measured in middle gluteal muscle samples. Postprandial insulin and glucose response was higher for diet A versus D. Log CK activity was higher with diets A, B, and C versus D. Daily insulin was higher and FFA lower on diet A versus B, C, and D, whereas glucose varied only slightly with diet. Muscle oxidative capacity and lipid stores were low in PSSM horses and muscle glycogen and abnormal polysaccharide content high on both diets A and D. Individual variation occurred in the response of PSSM horses to diets differing in starch and fat content. However, for those horses with clinical manifestations of PSSM, a diet with <5% DE starch and >12% DE fat can reduce exertional rhabdomyolysis, potentially by increasing availability of FFA for muscle metabolism.

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This study was designed to determine the effects of fat adaptation on carbohydrate and fat oxidation in conditioned horses during low-intensity exercise. Five mature Arabians were studied. The study was conducted as a crossover design with 2 dietary periods, each of 10 week's duration: a) a control (CON) diet, and b) a fat-supplemented (FAT) diet. The total amount of digestible energy (DE) supplied by the fat in the CON and FAT diets was 7% and 29%, respectively. During each period, the horses completed exercise tests at the beginning of the period (Week 0) and after 5 and 10 weeks on the diet. Tests consisted of 90 min of exercise at a speed calculated to elicit 35% VO2max on a treadmill inclined to 3 degrees. Oxygen consumption (VO2), carbon dioxide production (VCO2), and respiratory exchange ratio (RER) were measured at 15-min intervals. For determination of glucose kinetics, a stable isotope ([6-6-d2] glucose) technique was used. Compared to the CON diet, FAT diet consumption for 5-10 weeks was associated with an altered metabolic response to low-intensity exercise, as evidenced by a more than 30% reduction in the production and utilisation of glucose; a decrease in RER; a decrease in the estimated rate of whole-body carbohydrate utilisation; and an increase in the whole-body rate of lipid oxidation during exercise.

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The purpose of this study was to examine the effects of an external nasal strip (NS), frusemide (FR) and a combination of the 2 treatments (NS + FR) on exercise-induced pulmonary haemorrhage (EIPH) in Thoroughbred horses. It was hypothesised that both the NS and FR would attenuate EIPH as assessed by red blood cell count in bronchoalveolar lavage fluid. In random order, 8 horses completed each of 4 sprint exercise tests on a treadmill: 1) NS; 2) FR (0.5 mg/kg bwt i.v., 4 h pre-exercise); 3) NS + FR; and 4) control (C; no treatment). After a 5 min warm-up (4.5 m/s), horses completed 2 min running at 120% maximum oxygen consumption (VO2max) with the treadmill set at 3 degrees incline. Mean +/- s.d. running speed was 14.2+/-0.2 m/s. In the FR and NS + FR trials, horses carried weight equal to that lost as a result of frusemide administration. During exercise at 120% Vo2max, oxygen consumption (Vo2) and carbon dioxide production (Vco2) were measured at 15 s intervals. Plasma lactate concentration was measured in samples collected before exercise, at the end of the sprint and after 5 min cool-down at the trot. Thirty minutes after the run, bronchoalveolar lavage (BAL) was performed and the red cell count in the fluid quantified. Vo2 and Vco2 were significantly lower in NS and NS + FR trials than in the C and FR trials at the end of the sprint exercise protocol. However, plasma lactate concentrations did not differ among treatments. Compared with the C trial (61.1+/-30.5 x 10(6) red blood cells/ml BAL fluid), pulmonary haemorrhage was significantly (P<0.05) decreased in both the NS (15.9+/-4.0 x 106 RBC/ml) and FR (12.2+/-5.8 x 10(6) RBC/ml) trials. EIPH in the NS + FR trial (7.9+/-1.0 x 10(6) RBC/ml) was further diminished (P<0.05) compared to the NS trial, but not different from the FR trial. We conclude that both the external nasal strip and frusemide attenuate pulmonary haemorrhage in Thoroughbred horses during high-speed sprint exercise. The external nasal strip appears to lower the metabolic cost of supramaximal exertion in horses. Given the purported ergogenic effects of frusemide, the external nasal strip is a valuable alternative for the attenuation of EIPH.

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OBJECTIVE: To determine the effects of 3 rations (low grain, fat, high grain) on plasma creatine kinase (CK) activity and lactate concentration in Thoroughbred horses with recurrent exertional rhabdomyolysis (RER). ANIMALS: 5 Thoroughbreds with RER and 3 healthy Thoroughbreds (control horses). PROCEDURES: Rations were formulated to meet (low-grain and fat rations) or exceed (high-grain ration) daily energy requirements. Each ration was fed to horses in a crossover design for 3 weeks. Horses were exercised on a treadmill Monday through Friday; maximum speed on Monday and Friday was 11 m/s (6% slope), on Tuesday and Thursday was 9 m/s, and on Wednesday was 4.5 m/s. Plasma CK activity and lactate concentration were determined before and after exercise. RESULTS: Horses with RER fed the high-grain ration had significantly greater CK activity and change in CK activity 4 hours after exercise, compared with those fed the low-grain ration. Horses with RER exercised at the trot or canter had significantly greater increases in CK activity, compared with those exercised at the gallop. Plasma lactate concentrations after exercise were similar in control and affected horses. Lactate concentration and CK activity were not correlated in horses with RER. CONCLUSIONS AND CLINICAL RELEVANCE: Rations high in grain and formulated to exceed daily energy requirements may increase episodes of rhabdomyolysis in thoroughbred horses susceptible to RER.

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To delay the onset of fatigue, endurance horses are often fed at rest stops during races. The resulting increase in blood insulin may adversely inhibit lipolysis. In humans, ingestion of fructose produces a smaller insulin rise than glucose. This study compared glucose and fructose as carbohydrate supplements for endurance horses. Three Arabian geldings were given 300 g of fructose (F), glucose (G) or 50% glucose: 50% fructose (GF), in 1.5 L water, by stomach tube. In the Resting Test, carbohydrate was administered at rest. Following treatment, blood samples were taken every 30 min for 8 h, and feces were collected for 24 h. Treatment did not affect fecal weight or water content. Plasma glucose and insulin responses did not differ among treatments. Post-treatment (60 min), plasma L-lactate tended to be higher (P = 0.06) after the F and GF treatments than after the G treatment. In the Exercise Test, two treadmill exercise bouts at 0 degrees incline (Bout 1: 90 min; Bout 2: 120 min) were separated by a 1-h rest period. A total distance of 36.84 km was covered at a mean speed of 2.9 m/s. Carbohydrate was administered 45 min before Bout 2. Plasma glucose and insulin at the start of Bout 2 were higher (P = 0.02 and 0.03, respectively) with the GF treatment than with the F treatment. However, during exercise, plasma glucose concentrations did not differ among treatments. We conclude that fructose is well-absorbed by horses and rapidly converted to glucose.

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There is considerable debate among horsemen about how to feed horses before exercise. Should horses be fed or fasted before work and when should hay be fed relative to grain and/or exercise? Three experiments were conducted to evaluate if feeding hay with and without grain affects glycaemic and haematological responses in Thoroughbred (TB) horses at rest and during a simulated competition exercise test (CET) on a high-speed treadmill. In Experiment 1, 6 TB horses were fed hay at 3 different times relative to a grain meal. Time of feeding hay affected glycaemic response, plasma protein and water intake post grain feeding. During Experiment 2, 4 TB horses were used in a 4 x 4 Latin square design to determine whether feeding grain with or without hay prior to a CET would affect substrate utilisation and exercise. Feeding grain reduced free fatty acid (FFA) availability and increased blood glucose disappearance during exercise (P < 0.05). Feeding hay either along with grain or ad libitum the night before exercise resulted in reduced plasma volume (P < 0.05) and higher lactate production (P < 0.05) and heart rates (P < 0.05) during exercise. During Experiment 3, 4 TB horses were used in a 4 x 4 Latin square design experiment to determine whether feeding forage but no grain prior to CET would affect substrate utilisation and performance. Feeding only forage before exercise did not adversely affect performance. It was concluded that grain should be withheld from horses before exercise, but that small quantities of hay should be fed to ensure proper gastrointestinal tract function.

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Previous studies have associated recurrent exertional rhabdomyolysis (RER) with a diet high in soluble carbohydrate (CHO). The purpose of this study was to investigate the effect of 3 diets on clinical and metabolic parameters in 5 Thoroughbred horses with RER and 3 healthy Thoroughbreds performing a standardised exercise test (SET). Two diets were formulated to meet energy requirements for the amount of exercise being performed in the form of CHO or fat (21.4 Mcal DE/day). The third diet was formulated to provide 135% of the DE of the other 2 diets in the form of an excessive amount of carbohydrate (28.8 Mcal DE/day). Diets were fed in a crossover design for 3 week blocks and then horses performed a near maximal SET. Changes in heart rate (HR), plasma lactate, plasma glucose, total plasma solids, packed cell volume (PCV), muscle lactate and muscle glycogen concentration were measured immediately prior to, during, and 5 min after exercise. Serum creatine kinase (CK) activity was measured prior to and 4 h post SET. A 2-way ANOVA was used to examine the effect of group and dietary treatment. When dietary treatments were compared, horses fed the high-CHO diet had a mean pre-SET PCV and pre-SET HR that was higher than horses fed the fat diet (P = 0.06 and P = 0.07, respectively). Pre-SET heart rates were highest in RER horses consuming the high-CHO diet compared to RER horses consuming the low-CHO and fat diets (P = 0.02). Horses with RER had 4 h post SET CK activity greater than 400 u/l in 7/14 (50%) measurements compared to control horses which had CK activity greater than 400 u/l in 2/7 (29%) measurements. This study did not demonstrate a significant effect of diet on rhabdomyolysis, indicated by CK activity, or on the metabolic response to exercise. However, diet may have a calming effect on Thoroughbred horses with RER as manifested by decreased pre-exercise heart rates and decreased pre-exercise PCV in horses fed the fat diet.

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This study looked at the effect of feeding diets supplemented with either a predominantly saturated or unsaturated vegetable oil over a prolonged period to exercising horses. Eight Thoroughbred horses were assigned to 2 diet treatments and for 10 months were fed Timothy hay and oats, together with a fortified sweet feed supplemented with either a predominantly unsaturated (Un) or a saturated (S) vegetable oil so that approximately 19% DE (Digestible Energy) came from dietary fat and approximately 12% from either the Un or S source (AC). An increased amount of Un or S fortified sweet feed, replacing the oats, was then fed for a further 6 months (HF) so that approximately 27% DE came from fat and approximately 20% from the Un or S vegetable oil. Standardised incremental treadmill exercise (8-12 m/s) tests (STEP) and duplicate oral glucose tolerance tests (TOL) were carried out after 3, 6 and 9 months of the AC diet and after 3 and 6 months on the HF diet. There was no significant effect of dietary treatment or when the tests were undertaken (time) on the insulin or lactate responses to the STEP tests. Overall there was a significant (P < 0.05) effect of time and treatment on the glucose response, but there was no difference between treatments at the first and last tests or between the results for these tests or between the endAC and endHF tests. No significant effect of treatment or time was seen on the TOL glucose response (% change from Time '0') although there was a trend for the glucose concentrations to be lower and the insulin responses higher (nonsignificant) in the S treatment group. No significant effect of treatment on haematological parameters, monitored monthly, was found. Total protein and gamma glutamyl transferase remained within the normal range throughout. There was a significant effect of treatment (P < 0.05) on cholesterol and triglycerides with higher concentrations in the S group from the first (1 month) sample. Linoleic acid was the main fatty acid in all the 4 plasma lipid classes with slightly, but significant (P < 0.05), higher concentrations in Un for the cholesterol ester and phospholipid classes. There was no effect of time. Overall, the total resting plasma fatty acid content was significantly higher (P < 0.05) with S at the sample points (endAC and endHF). No adverse effects of feeding either diet on apparent coat condition or hoof appearance were seen apart from an apparent increase in the grease score. Many of the parameters assessed showed significant improvements with time (P < 0.05). In conclusion, no apparent adverse effects of feeding a diet supplemented with either an unsaturated or saturated vegetable oil for 6 months at approximately 20% DE after 10 months at approximately 12% DE were identified and there were no apparent disadvantages of feeding a saturated vegetable oil supplemented diet compared with an unsaturated one.

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Six Thoroughbreds were used to evaluate time of feeding on changes in exercise response in horses receiving either a textured feed or a fat-supplemented textured feed. Using a crossover design, 3 horses were fed a fat-supplemented diet while 3 horses received a control ration of textured feed. Horses performed a standardised exercise test (SET) on a high speed treadmill. The SET was performed at 3 different times: 1) following an overnight 12 h fast, 2) 3 h after feeding and 3) 8 h after feeding. The SET consisted of a 2 min walk at 1.4 m/s, 800 m trot at 4.2 m/s, 800 m gallop at 7.7 m/s, 1600 m gallop at 11 m/s, 800 m trot at 4.2 m/s and 2 min walk at 1.4 m/s. Jugular blood samples were taken before feeding, hourly until the beginning of the SET, at the end of each exercise step, 15 min post exercise and 30 min post exercise. During the SET, heart rate was measured and blood samples collected for analysis of glucose, lactate, insulin and nonesterified fatty acids (NEFA). Feeding horses 3 h prior to exercise resulted in elevated concentrations of plasma glucose and insulin (P < 0.01) at rest. Elevated concentrations of insulin in horses fed 3 h prior to exercise decreased plasma glucose (P < 0.01) during exercise and appeared to have suppressed fat oxidation during exercise because horses that were either fasted or fed 8 h post prandial had a net disappearance of NEFA in the plasma during exercise. This study indicates that beginning exercise with elevated plasma insulin appeared to be of no benefit during the exercise conducted in this experiment.

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High-grain diets for the exercising horse were compared with diets which provided 15% of the total caloric intake from either vegetable oil or a highly fermentable fibre source (beet pulp). Six Thoroughbreds age 3 years were fed one of 3 diets or 5 weeks in a replicated 3 x 3 Latin square. The CONTROL diet was 3.65 kg of sweet feed (SF), 0.9 kg of a protein/vitamin/mineral pellet and 5.45 kg of hay cubes. The FAT diet replaced 1.15 kg of SF with 0.45 kg of soybean oil and the FIBRE diet replaced 1.15 kg of SF with 1.36 kg of beet pulp. Horses were exercised 3 times per week on a high-speed treadmill. During the last week of each period, the horses performed a standardised exercise test (SET). A series of blood samples was drawn immediately before feeding and every 0.5 h for 3 h after feeding, throughout the exercise bout and 30 min post exercise. Plasma was analysed for lactate, glucose, cortisol, insulin, packed cell volume, total protein and triglycerides. Water intake was measured at regular intervals during SET day. Blood glucose was lower (P < 0.05) in the FAT-fed horses during the 3 h post feeding as compared to either CONTROL or FIBRE-fed horses. Insulin was lower (P < 0.05) in the FAT-fed both post feeding and throughout exercise. Cortisol was lower (P < 0.05) in the FAT than the CONTROL-fed during exercise. Following exercise, the FAT-fed drank more water (P < 0.01) than either CONTROL or FIBRE-fed. Substituting 15% of DE as vegetable oil had a greater effect on metabolic response to exercise than a 15% substitution of beet pulp.

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Energy balance studies using indirect calorimetry were conducted with four mature equids ranging in weight from 125 to 856 kg. Each animal was fed three different levels of intake of the same diet. The amounts of digestible and metabolizable energy required for zero energy balance were determined by calculating regression equations for energy balance against energy intake. It was concluded that the maintenance requirements of equids vary linearly with body weight. No advantage was found for the use of weight for the comparison of equids within the range studied. The digestible energy (DE) requirement for equids confined to metabolism stalls was DE (kcal/d) = 975 + 21.28 X W where W equals body weight in kg. If a factor for activity similar to that suggested by the National Research Council was included, the requirement could be calculated by the equation: DE (kcal/d) = 1,375 + 30.0 X W.

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Energy expenditure was measured in four geldings (433 to 520 kg) during submaximal exercise on a racetrack using a mobile open-circuit indirect respiration calorimeter. A total of 304 5-min measurements of O2 consumed and CO2 produced were taken. Measurements were made with and without riders. The amount of energy expended by the horses was exponentially related to speed and was proportional to the body weight of the riderless horse or the combined weight of the horse plus rider and tack. Total energy expended by the four horses walking, trotting, cantering was best described by the equation: Y = e3.02 + .0065X where Y = energy expended (cal X kg-1 X min-1) and X = speed (m/min). Digestible energy (DE) required above maintenance was calculated as (Formula: see text).

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Twenty-two pony mares were fed one of three diets that provided 93.0, 74.8 or 57.2 kcal of digestible energy (DE) per kg body weight per day. Milk samples were taken at 14 day intervals. A total of five samples were taken from each mare. The samples were analyzed for total solids, crude protein, lactose, total lipids, ash, calcium and phosphorus. Gross energy was calculated from composition data. Increases in energy intake decreased the concentration of total solids, protein, fat and gross energy of mare's milk. Energy intake had a greater influence on the mare's body condition than on milk energy production. It was concluded the objective of a feeding program for a lactating mare should be to keep the mare in a desirable body condition rather than to influence milk composition or production.

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Forty-eight mutants unable to reduce nitrate were isolated from "cowpea" Rhizobium sp. strain 32Hl and examined for nitrogenase activity in culture. All but two of the mutants had nitrogenase activity comparable with the parental sttain and two nitrogenase-defective strains showed alterations in their symbiotic properties. One strain was unable to nodulate either Macroptilium atropurpureum or Vigna uguiculata and, with the other, nodules appeared promptly, but effective nitrogen fixation was delayed. These results, and the relatively low proportion of nitrate reductase mutants with impaired nitrogenase activity, do not support the proposed commanality between nitrogenase and nitrate reductase in cowpea rhizobia. Inhibition studies of the effect of nitrate and its reduction products on the nitrogenase activity in cultured strains 32Hl and the nitrate reductase-deficient, Nif+ strains, indicated that nitrogenase activity was sensitive to nitrite rather than to nitrate.

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The effect of nitrate on the symbiotic properties of nitrate-reductase-deficient mutants of a strain of cowpea rhizobia (32H1), and of a strain of Rhizobium trifolii (TA1), were examined; the host species were Macroptilium atropurpureum (DC.) Urb. and Trifolium subterraneum L. Nitrate retarded initial nodulation by the mutant strains to an extent similar to that found with the parent strains. It is therefore unlikely that nitrite produced from nitrate by the rhizobia, plays a significant role in the inhibition of nodulation by nitrate. Nitrite is an inhibitor of nitrogenase, and its possible production in the nodule tissue by the action of nitrate reductase could be responsible for the observed inhibition of nitrogen fixation when nodulated plants are exposed to nitrate. However, the results of this investigation show that nitrogen fixation by the plants nodulated by parent or mutant strains was depressed by similar amounts in the presence of nitrate. No nitrite was detected in the nodules. Nodule growth, and to a lesser extent, the nitrogenase specific activity of the nodules (µmol C2H4g(-1) nodule fr. wt. h(-1)), were both affected by the added nitrate.

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Nutritional and physical conditions affecting nitrogenase activity in the strain of "cowpea" rhizobia, 32H1, were examined using cultures grown on agar medium. Arabinose in the basic medium (CS7) could be replaced by ribose, xylose, or glycerol, but mannitol, glucose, sucrose, or galactose only supported low nitrogenase (C2H2 reduction) activity. Succinate could be replaced by pyruvate, fumarate, malate, or 2-oxoglutarate, but without any carboxylic acid, nitrogenase activity was low or undetectable unless a high level of arabinose was provided. Inositol was not essential. Several nitrogen sources could replace glutamine including glutamate, urea, (NH4)2SO4 and asparagine. The maximum nitrogenase activity of cultures grown in air at 30 degrees C was observed under assay conditions of pO2=0.20-0.25 atm and 30 degrees C incubation. Greatest activity occurred after a period of rapid bacterial growth, when viable cell count was relatively constant. Compared with results obtained on the CS7 medium, nitrogenase activity could be substantially increased and/or sustained for longer periods of time by using 12.5 MM succinate and 100 mM arabinose, by increasing phosphate concentration from 2 to 30-50 mM, or by culturing the bacteria at 25 degrees C.

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Nitrogen fixation was induced in a strain of "cowpea" rhizobia, 32Hl, when it was grown in association with cell cultures of the non-legume, tobacco (Nicotiana tabacum). Rhizobia grown alone on the various media examined did not show nitrogenase activity, indicating the involvement of particular plant metabolites in nitrogenase induction. Nitrogenase activity, as measured by C2H2 reduction, was maximized at an O2 concentration of 20% and at an assay temperature of 30°C, the conditions under which the plant cell-rhizobia associations developed. Glutamine, as a nitrogen source, could be replaced by other organic nitrogen sources, but NH4 (+) and NO3 (-) repressed nitrogenase activity. Nitrogenase activity induced in rhizobia when cultured adjacent to, but not in contact with, the plant cells could be stimulated by providing succinate in the medium. At least 12 other strains of rhizobia also reduced C2H2 in association with tobacco cells; the highest levels of activity were found among cowpea strains.

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